Abstract
DNA methylation alterations are early events during tumourigenesis, affecting genes involved in the crosstalk between cells and surroundings in colorectal cancer (CRC). Among these genes, GRIA4, Glutamate Ionotropic Receptor AMPA Type Subunit 4, displays hypermethylation in the promoter region, and is an early diagnostic biomarker. It is well known that methylation can also affect alternative transcription. The purpose of this study is to evaluate the expression, at transcript and protein level, of GRIA4 main isoforms (the canonical one and a short variant) in 23 CRC and matched normal samples, of which we previously verified the methylation status. We further predicted miRNA/transcript target interactions as a possible post-transcriptional regulation using bioinformatics tools. As expected, downregulation of both variants has been observed in tumours. Interestingly, in contrast to what observed at transcriptional level, the GluR4 protein short isoform displayed higher expression than the canonical one either in normal or tumoural tissues. This may be explained by miRNA specifically targeting the canonical isoform. Our study is the first one that shows the expression of both isoforms in colon tissues. To note, the evident expression of the short isoform suggests a functional role in intestinal cell biology.
Highlights
Cancer cells are characterized by massive changes in gene expression profile in terms of transcripts levels and expression of alternative isoforms, due to different biological mechanisms, among which alterations in DNA methylation patterns have a pivotal role [1,2,3,4,5,6,7,8].DNA methylation at cytosines predominantly occurs in CpG dinucleotides known as CpG sites
In a genome-wide methylation study, we identified 74 CpG islands (CGIs) significantly aberrantly methylated shared between colorectal cancer (CRC) and adenoma samples
We evaluated the expression levels, using qRT-PCR and Western blot, of selected genes, barely expressed in normal colon, whose promoter CGIs were hypermethylated in CRC and we detected their further downregulation in tumour samples [1, 3]
Summary
Cancer cells are characterized by massive changes in gene expression profile in terms of transcripts levels and expression of alternative isoforms, due to different biological mechanisms, among which alterations in DNA methylation patterns have a pivotal role [1,2,3,4,5,6,7,8].DNA methylation at cytosines predominantly occurs in CpG dinucleotides known as CpG sites. Cancer cells are characterized by massive changes in gene expression profile in terms of transcripts levels and expression of alternative isoforms, due to different biological mechanisms, among which alterations in DNA methylation patterns have a pivotal role [1,2,3,4,5,6,7,8]. DNA methylation dramatically changes in cancer cells, i.e. a wide loss of DNA methylation occurs, whereas promoter-associated CGIs, usually un-methylated, undergo de novo methylation [4]. These aberrations, which can be cancer-specific, are considered early and frequent events in tumourigenesis, sometimes detected in premalignant tissues, becoming promising diagnostic biomarkers [9]. DNA methylation alterations can be traced in cell-free circulating tumour DNA through non-invasive techniques [1, 3, 11, 12]
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